Amide exchange shows calcium-induced conformational changes are transmitted to the dimer interface of S100B.

S100B is a 21 kDa member of the S100 calcium-binding protein family. This protein comprises a symmetric homodimer with each subunit having two EF-hands arranged from four alpha-helices (I-IV). S100B binds calcium and undergoes a conformation change leading to the exposure of hydrophobic surface residues that enable the protein to interact with biological target molecules. The most significant structural change that occurs during calcium binding results in a change in the orientation of helix III with respect to helices II and IV. In this work, the calcium-sensitive conformational change has been studied by utilizing fast 1H-15N HSQC experiments and water-transfer methods to follow the amide exchange in apo-S100B and Ca-S100B at 35 degrees C. In apo-S100B, the protection factors are 2-3 orders of magnitude lower for helix III than for helix I, II, or IV. In addition, the exchange stability measured here for the dimer interface helices (I, I', IV, and IV'), in the absence of calcium, is similar to the stability obtained from chemical denaturation experiments. When calcium binds, significant decreases in the protection factors for helices I and IV indicate a modification in the stability of the dimer interface has occurred. In contrast, helix II protection factors increase slightly, which is consistent with a decreased level of surface exposure of this helix. These data have been compared with those of the monomeric S100 protein, calbindin D9k, to illustrate that upon calcium binding there is a balance maintained between the amide exchange rates in helices II and III, although largely the rates are dissimilar for each of these proteins. This distinguishing feature may be important for the calcium-induced conformational change in S100B, where calcium binding is transmitted to the dimer-forming helices.